Presence and fate of antibiotic residues, antibiotic resistance genes and zoonotic bacteria during biological swine manure treatment.

The presence and dissemination of antibiotic residues, antibiotic resistance genes and zoonotic bacteria in the environment is of growing concern worldwide. Manure management practices, such as biological removal of nitrogen from swine manure, may help to decrease levels of antibiotic residues, antibiotic resistance genes and zoonotic bacteria present in manure before fertilization, thereby reducing environmental contamination. Therefore, the aim of this study was to monitor the presence and fate of seven antibiotic residues (colistin, sulfadiazine, trimethoprim, doxycycline, oxytetracycline, ceftiofur and tylosin A), nine antibiotic resistance genes (tet(B), tet(L), tet(M), tet(O), tet(Q), tet(W), erm(B), erm(F) and sul2) and two zoonotic bacteria (Salmonella Typhimurium and Campylobacter coli) during biological nitrogen removal from swine manure over time. Samples from the raw manure, the solid fraction, the liquid fraction and the storage lagoon were analyzed on two farms at six time points with an interval of two weeks. Only the antibiotics which were used during the three months preceding the first sampling could be detected before and after biological nitrogen removal from swine manure. Of all the antibiotics studied, doxycycline was recovered in all of the samples and sulfadiazine was recovered in most samples on both farms. For both antibiotics, there appears to be a reduction of the amount of residues present in the storage lagoon compared to the liquid fraction, however, this reduction was not statistically significant. A significant reduction of the relative abundances of most of the antibiotic resistance genes studied was observed when comparing the liquid fraction and the storage lagoon. For tet(L), no differences were observed between the fractions sampled and for sul2 and erm(F), a significant increase in relative abundances was observed on the second farm sampled. For the zoonotic bacteria, a reduction of at least 1 log was observed after biological nitrogen removal from swine manure. The results indicate that the concentration of certain antibiotic residues and several antibiotic resistance genes and the amount of zoonotic bacteria present in the manure may be reduced in the end product of the biological nitrogen removal from swine manure.

EFSA's scientific activities and achievements on the risk assessment of genetically modified organisms (GMOs) during its first decade of existence: looking back and ahead.

Genetically modified organisms (GMOs) and derived food and feed products are subject to a risk analysis and regulatory approval before they can enter the market in the European Union (EU). In this risk analysis process, the role of the European Food Safety Authority (EFSA), which was created in 2002 in response to multiple food crises, is to independently assess and provide scientific advice to risk managers on any possible risks that the use of GMOs may pose to human and animal health and the environment. EFSA's scientific advice is elaborated by its GMO Panel with the scientific support of several working groups and EFSA's GMO Unit. This review presents EFSA's scientific activities and highlights its achievements on the risk assessment of GMOs for the first 10 years of its existence. Since 2002, EFSA has issued 69 scientific opinions on genetically modified (GM) plant market registration applications, of which 62 for import and processing for food and feed uses, six for cultivation and one for the use of pollen (as or in food), and 19 scientific opinions on applications for marketing products made with GM microorganisms. Several guidelines for the risk assessment of GM plants, GM microorganisms and GM animals, as well as on specific issues such as post-market environmental monitoring (PMEM) were elaborated. EFSA also provided scientific advice upon request of the European Commission on safeguard clause and emergency measures invoked by EU Member States, annual PMEM reports, the potential risks of new biotechnology-based plant breeding techniques, evaluations of previously assessed GMOs in the light of new scientific publications, and the use of antibiotic resistance marker genes in GM plants. Future challenges relevant to the risk assessment of GMOs are discussed. EFSA's risk assessments of GMO applications ensure that data are analysed and presented in a way that facilitates scientifically sound decisions that protect human and animal health and the environment.

Genetic diversity of livestock-associated MRSA isolates obtained from piglets from farrowing until slaughter age on four farrow-to-finish farms.

During a previous longitudinal study, performed on four farrow-to-finish farms (A to D), samples were taken from twelve sows, their offspring, and the environment on various occasions over six months to study the MRSA presence. During the present study, a selection of the obtained MRSA isolates were typed by multiple-locus variable-number tandem-repeat analysis (MLVA), Pulsed Field Gel Electrophoresis (PFGE), spa typing, and SCCmec typing to study the genetic diversity of LA-MRSA isolates and to determine possible MRSA sources for pig(let)s. PFGE, spa typing, and SCCmec typing revealed the presence of one or few dominant genotype(s) per farm. In contrast, 212 MLVA types were detected on the four farms, forming one cluster on farm A, three on farm B, four on farm C and two on farm D. The genotype, found on farm A was unique for this farm. Farms B, C and D shared one cluster. In general, MLVA types from these clusters were isolated from piglets, sows, and the environment on various sampling events. Piglets carried MLVA types both related and unrelated to their mother sows' MLVA types at farrowing and onwards. In conclusion, molecular typing revealed that within a farm one or a few dominant strain(s) are widespread. Potential MRSA sources for piglets were mother sows, the environment and other piglets.

Safety and efficacy of a feed additive consisting of l-tryptophan (produced with Escherichia coliCGMCC 7.460) for all animal species (Kempex Holland B.V.).

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of the feed additive consisting of l-tryptophan produced by fermentation with Escherichia coli CGMCC 7.460 when used as a nutritional additive in feed and water for drinking for all animal species and categories. The production strain is not genetically modified. Viable cells of the production strain were not detected in the final additive. The additive does not give rise to any safety concern regarding the production strain. The use of l-tryptophan (≥ 98%) produced with E. coli CGMCC 7.460 to supplement feed is safe for non-ruminant species. There may be a risk for an increased production of toxic metabolites when unprotected tryptophan is used in ruminants. The EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) has concerns on the safety of the simultaneous oral administration of l-tryptophan via water for drinking and feed due to possible amino acid imbalances and hygienic reasons. The use of l-tryptophan produced with E. coli CGMCC 7.460 in animal nutrition raises no safety concerns to consumers of animal products and to the environment. In the absence of data, the FEEDAP Panel cannot conclude on the potential of the additive to be irritant to skin or eyes, or on its potential to be a dermal sensitiser. The endotoxin activity of the additive in combination with the high dusting potential may represent a risk of exposure by inhalation to endotoxins for users. The additive l-tryptophan is regarded as an effective source of the amino acid l-tryptophan for all non-ruminant species. To be as efficacious in ruminants as in non-ruminants, it should be protected from ruminal degradation.

Evaluation of alternative methods of tunnel composting (submitted by the European Composting Network) II.

Two alternative methods for producing compost in a tunnel, from certain category (Cat.) 3 animal by-products (ABP) and other non-ABP material, were assessed. The first method proposed a minimum temperature of 55°C for 72 h and the second 60°C for 48 h, both with a maximum particle size of 200 mm. The assessment of the Panel on Biological Hazards (BIOHAZ) exclusively focused on Cat. 3 ABP materials (catering waste and processed foodstuffs of animal origin no longer intended for human consumption). The proposed composting processes were evaluated for their efficacy to achieve a reduction of at least 5 log10 of Enterococcus faecalis and Salmonella Senftenberg (775W, H2S negative) and at least 3 log10 of relevant thermoresistant viruses. The applicant provided a list of biological hazards that may enter the composting process and selected parvoviruses as the indicator of the thermoresistant viruses. The evidence provided by the applicant included: (a) literature data on thermal inactivation of biological hazards; (b) results from validation studies on the reduction of E. faecalis, Salmonella Senftenberg 775W H2S negative and canine parvovirus carried out in composting plants across Europe; (c) and experimental data from direct measurements of reduction of infectivity of murine parvovirus in compost material applying the time/temperature conditions of the two alternative methods. The evidence provided showed the capacity of the proposed alternative methods to reduce E. faecalis and Salmonella Senftenberg 775W H2S negative by at least 5 log10, and parvoviruses by at least 3 log10. The BIOHAZ Panel concluded that the two alternative methods under assessment can be considered to be equivalent to the processing method currently approved in the Commission Regulation (EU) No 142/2011.

Scientific opinion on the characterisation of Ensifer adhaerens CGMCC 19596 used in the production of the feed additive consisting of vitamin B12 (cyanocobalamin) for all animal species (Hebei Huarong Pharmaceutical Co., ltd.).

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the characterisation of the feed additive consisting of vitamin B12 (cyanocobalamin) produced by fermentation with Ensifer adhaerens (CGMCC 19596). The additive is intended to be used as a nutritional additive for all animal species. In a previous opinion, the FEEDAP Panel could not conclude on the characterisation of the production strain, due to uncertainties on whether the production strain E. adhaerens CGMCC 19596 was genetically modified. However, since viable cells and DNA were not detected in the product, the FEEDAP Panel concluded that vitamin B12 (cyanocobalamin), produced with E. adhaerens CGMCC 19596 would not raise safety concerns as regards the production strain. In the present submission, the applicant provided supplementary information regarding the origin and history of modifications of the strain. Based on the data provided, the FEEDAP Panel concluded on the characterisation of the production strain E. adhaerens CGMCC 19596, which can be considered not to be genetically modified.

BSE risk posed by ruminant collagen and gelatine derived from bones.

The European Commission requested an estimation of the BSE risk (C-, L- and H-BSE) from gelatine and collagen derived from ovine, caprine or bovine bones, and produced in accordance with Regulation (EC) No 853/2004, or Regulation (EC) No 1069/2009 and its implementing Regulation (EU) No 142/2011. A quantitative risk assessment was developed to estimate the BSE infectivity, measured in cattle oral infectious dose 50 (CoID50), in a small size batch of gelatine including one BSE-infected bovine or ovine animal at the clinical stage. The model was built on a scenario where all ruminant bones could be used for the production of gelatine and high-infectivity tissues remained attached to the skull (brain) and vertebral column (spinal cord). The risk and exposure pathways defined for humans and animals, respectively, were identified. Exposure routes other than oral via food and feed were considered and discussed but not assessed quantitatively. Other aspects were also considered as integrating evidence, like the epidemiological situation of the disease, the species barrier, the susceptibility of species to BSE and the assumption of an exponential dose-response relationship to determine the probability of BSE infection in ruminants. Exposure to infectivity in humans cannot be directly translated to risk of disease because the transmission barrier has not yet been quantified, although it is considered to be substantial, i.e. much greater amounts of infectivity would be needed to successfully infect a human and greater in the oral than in the parenteral route of exposure. The probability that no new case of BSE in the cattle or small ruminant population would be generated through oral exposure to gelatine made of ruminant bones is 99%-100% (almost certain) This conclusion is based on the current state of knowledge, the epidemiological situation of the disease and the current practices, and is also valid for collagen.

Modification of the terms of authorisation regarding the additive consisting of liquid l-lysine base produced with Corynebacterium glutamicum NRRL B-67439 and NRRL B-67535 for all animal species (ADM specialty ingredients (Europe) B.V.).

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of liquid l-lysine base produced with a genetically modified strain of Corynebacterium glutamicum as a nutritional feed additive for all animal species. The l-lysine base liquid produced with C. glutamicum NRRL B-67535 and NRRL B-67439 is currently authorised as a nutritional additive for all animal species. The present application is aimed at modifying the current authorisation to include C. glutamicum NRRL B-68248 as a production strain. The new production strain qualifies for the qualified presumption of safety approach when used for production purposes. It was unambiguously identified as C. glutamicum and was shown not to harbour acquired antimicrobial resistance determinants for antibiotics of human and veterinary importance. All the introduced sequences or mutations were considered to be safe, and no viable cells or DNA of the NRRL B-68248 strain was detected in the final product. Therefore, the final product does not pose any safety concern associated with the production strain. l-Lysine base produced using C. glutamicum NRRL B-68248 does not represent a risk for the target species, the consumer or the environment. The additive was considered to be neither irritant to skin or the eyes, nor a dermal sensitiser. l-Lysine base produced with C. glutamicum NRRL B-68248 is considered to be an efficacious source of the essential amino acid l-lysine for non-ruminant animal species. For the supplemental l-lysine to be as efficacious in ruminants as in non-ruminant species, it would require protection against degradation in the rumen.

Safety evaluation of the food enzyme mucorpepsin from the non-genetically modified Rhizomucor miehei strain LP-N836.

The food enzyme mucorpepsin (EC 3.4.23.23) is produced with the non-genetically modified Rhizomucor miehei strain LP-N836 by Meito Sangyo Co., Ltd. The native enzyme can be chemically modified to produce a more thermolabile form. The food enzyme is free from viable cells of the production organism. It is intended to be used in the processing of dairy products for the production of cheese and fermented dairy products. Dietary exposure to the food enzyme-total organic solids (TOS) was estimated to be up to 0.108 mg TOS/kg body weight (bw) per day in European populations. Genotoxicity tests did not indicate a safety concern. The systemic toxicity was assessed by means of a repeated dose 90-day oral toxicity study in rats. The Panel identified a no observed adverse effect level of 95 mg TOS/kg bw per day, the mid-dose tested, which when compared with the estimated dietary exposure, resulted in a margin of exposure of at least 880. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and four matches with respiratory allergens and one with a food allergen (mustard) were found. The Panel considered that the risk of allergic reactions upon dietary exposure to this food enzyme, particularly in individuals sensitised to mustard proteins, cannot be excluded. Based on the data provided, the Panel concluded that both the native and thermolabile forms of this food enzyme do not give rise to safety concerns under the intended conditions of use.

Safety evaluation of the food enzyme AMP deaminase from non-genetically modified Aspergillus sp. strain DEA 56-111.

The food enzyme AMP deaminase (AMP aminohydrolase; EC 3.5.4.6) is produced with the non-genetically modified microorganism Aspergillus sp. strain DEA 56-111 by Shin Nihon Chemical Co., Ltd. The food enzyme was considered free from viable cells of the production organism. It is intended to be used in the processing of yeast and yeast products. Dietary exposure to the food enzyme-total organic solids (TOS) was estimated to be up to 0.005 mg TOS/kg body weight (bw) per day in European populations. Genotoxicity tests did not indicate a safety concern. The Panel identified a no observed adverse effect level of 1984 mg TOS/kg bw per day, the highest dose tested, which, when compared with the estimated dietary exposure, resulted in a margin of exposure of at least 396,800. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that the risk of allergic reactions upon dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns, under the intended conditions of use.

Safety and efficacy of a feed additive consisting of inactivated selenised yeast (Saccharomyces cerevisiae CCTCC M 2022402) for all animal species (Phytobiotics Futterzusatzstoffe GmbH).

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of the selenised yeast (inactivated) Saccharomyces cerevisiae CCTCC M 2022402 (Plexomin® Se 3000, available in two forms: 'granules' and 'micro') as a nutritional feed additive for all animal species. Based on a tolerance-efficacy trial, the FEEDAP Panel concluded that the additive is safe for chickens for fattening at proposed conditions of use and this conclusion can be extrapolated to all animal species. In the absence of deposition data in all animal species and products, the FEEDAP Panel cannot conclude on the safety for the consumer. Plexomin® Se 3000 (granules) is dust-free; therefore, the exposure through inhalation is unlikely. Plexomin® Se 3000 (micro) presents a risk by inhalation. Both forms of the additive (granules and micro) are considered as respiratory sensitisers. Due to the lack of data, no conclusions can be drawn on the dermal and eye irritation potential of Plexomin® Se 3000 (granules). Plexomin® Se 3000 (micro) is not irritant to the skin and the eyes. No conclusions can be drawn on the potential of both forms of the additive to be dermal sensitisers. The use of the additive in animal nutrition is considered safe for the environment. The additive is an efficacious source of selenium in feedingstuffs for all animal species.

Safety evaluation of the food enzyme bacillolysin from the non-genetically modified Bacillus amyloliquefaciens strain NZYM-NB.

The food enzyme bacillolysin (EC 3.4.24.28) is produced with the non-genetically modified Bacillus amyloliquefaciens strain NZYM-NB by Novozymes A/S. The production strain meets the requirements for qualified presumption of safety (QPS) approach to safety assessment. The food enzyme is intended to be used in eleven food manufacturing processes. Since residual amounts of total organic solids (TOS) are removed during two processes, dietary exposure was estimated only for the remaining nine food manufacturing processes. Exposure was estimated to be up to 1.327 mg TOS/kg body weight per day in European populations. As the production strain qualifies for the QPS approach and no issue of concern arising from the production process of the food enzyme was identified, the Panel considered that no toxicological studies other than the assessment of allergenicity were necessary. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded (except for distilled alcohol production), but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.

Safety evaluation of the food enzyme microbial collagenase from the genetically modified Streptomyces violaceoruber strain pCol.

The food enzyme microbial collagenase (EC 3.4.24.3) is produced with the genetically modified Streptomyces violaceoruber strain pCol by Nagase (Europa) GmbH. The genetic modifications do not give rise to safety concerns. The food enzyme is free from viable cells of the production organism and its DNA. It is intended to be used in two food manufacturing processes: the production of modified meat and fish products and the production of protein hydrolysates from meat and fish proteins. The dietary exposure to the food enzyme-total organic solids (TOS) was estimated to be up to 1.098 mg TOS/kg body weight (bw) per day in European populations. Genotoxicity tests did not indicate a safety concern. The systemic toxicity was assessed by means of a repeated dose 90-day oral toxicity study in rats. The Panel identified a no observed adverse effect level of 940 mg TOS/kg bw per day, the highest dose tested, which, when compared with the estimated dietary exposure, resulted in a margin of exposure of at least 856. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns, under the intended conditions of use.

Safety evaluation of the food enzyme bacillolysin from the non-genetically modified Bacillus amyloliquefaciens strain DP-Cyb74.

The food enzyme bacillolysin (EC 3.4.24.28) is produced with the non-genetically modified Bacillus amyloliquefaciens strain DP-Cyb74 by Genencor International B.V. The production strain met all requirements for the qualified presumption of safety (QPS) approach to safety assessment. The food enzyme is intended to be used in six food manufacturing processes. Dietary exposure to the food enzyme total organic solids (TOS) was estimated to be up to 1.536 mg TOS/kg body weight per day in European populations. As the production strain qualifies for the QPS approach and no issue of concern arose from the production process of the food enzyme, the Panel considered that no toxicological studies other than the assessment of allergenicity were necessary. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.

Safety evaluation of the food enzyme thermolysin from the non-genetically modified Anoxybacillus caldiproteolyticus strain AE-TP.

The food enzyme thermolysin (EC. 3.4.24.27) is produced with the non-genetically modified Anoxybacillus caldiproteolyticus strain AE-TP by Amano Enzyme Inc. The food enzyme is free from viable cells of the production organism. It is intended to be used in eight food manufacturing processes. Dietary exposure was estimated to be up to 0.973 mg TOS/kg body weight (bw) per day in European populations. Genotoxicity tests did not indicate a safety concern. The systemic toxicity was assessed by means of a repeated dose 90-day oral toxicity study in rats. The Panel identified a no observed adverse effect level of 700 mg TOS/kg bw per day, the mid-dose tested, which, when compared with the estimated dietary exposure, resulted in a margin of exposure of at least 719. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no matches were found. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.

Safety evaluation of the food enzyme ß-fructofuranosidase from the non-genetically modified Saccharomyces cerevisiae strain NCYC R693.

The food enzyme ß-fructofuranosidase (ß-d-fructofuranoside fructohydrolase; EC 3.2.1.26) is produced with the non-genetically modified Saccharomyces cerevisiae strain NCYC R693 by Kerry Ingredients & Flavours Ltd. The production strain meets the requirements for the qualified presumption of safety (QPS) approach. The food enzyme is intended to be used in four food manufacturing processes. The dietary exposure to the food enzyme-total organic solids (TOS) was estimated to be up to 2.485 mg TOS/kg body weight per day in European populations. As the production strain qualifies for the QPS approach of safety assessment and no issue of concern arising from the production process of the food enzyme were identified, the Panel considered that no toxicological studies other than the assessment of allergenicity were necessary. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and one match with a tomato allergen was found. The Panel considered that the risk of allergic reactions upon dietary exposure to this food enzyme, particularly in individuals sensitised to tomato, cannot be excluded. However, the likelihood of allergic reactions is expected not to exceed the likelihood of allergic reactions to tomato. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.

Taxonomic identity of the Bacillus licheniformis strains used to produce food enzymes evaluated in published EFSA opinions.

Bacillus paralicheniformis, a species known to produce the antimicrobial bacitracin, could be misidentified as Bacillus licheniformis, depending on the identification method used. For this reason, the European Commission requested EFSA to review the taxonomic identification of formerly assessed B. licheniformis production strains. Following this request, EFSA retrieved the raw data from 27 technical dossiers submitted and found that the taxonomic identification was established by 16S rRNA gene analyses for 15 strains and by whole genome sequence analysis for 12 strains. As a conclusion, only these 12 strains could be unambiguously identified as B. licheniformis.

Re-evaluation of certain aspects of the EFSA Scientific Opinion of April 2010 on risk assessment of parasites in fishery products, based on new scientific data. Part 1: ToRs1-3.

Surveillance data published since 2010, although limited, showed that there is no evidence of zoonotic parasite infection in market quality Atlantic salmon, marine rainbow trout, gilthead seabream, turbot, meagre, Atlantic halibut, common carp and European catfish. No studies were found for greater amberjack, brown trout, African catfish, European eel and pikeperch. Anisakis pegreffii, A. simplex (s. s.) and Cryptocotyle lingua were found in European seabass, Atlantic bluefin tuna and/or cod, and Pseudamphistomum truncatum and Paracoenogonimus ovatus in tench, produced in open offshore cages or flow-through ponds or tanks. It is almost certain that fish produced in closed recirculating aquaculture systems (RAS) or flow-through facilities with filtered water intake and exclusively fed heat-treated feed are free of zoonotic parasites. Since the last EFSA opinion, the UV-press and artificial digestion methods have been developed into ISO standards to detect parasites in fish, while new UV-scanning, optical, molecular and OMICs technologies and methodologies have been developed for the detection, visualisation, isolation and/or identification of zoonotic parasites in fish. Freezing and heating continue to be the most efficient methods to kill parasites in fishery products. High-pressure processing may be suitable for some specific products. Pulsed electric field is a promising technology although further development is needed. Ultrasound treatments were not effective. Traditional dry salting of anchovies successfully inactivated Anisakis. Studies on other traditional processes - air-drying and double salting (brine salting plus dry salting) - suggest that anisakids are successfully inactivated, but more data covering these and other parasites in more fish species and products is required to determine if these processes are always effective. Marinade combinations with anchovies have not effectively inactivated anisakids. Natural products, essential oils and plant extracts, may kill parasites but safety and organoleptic data are lacking. Advanced processing techniques for intelligent gutting and trimming are being developed to remove parasites from fish.

Safety evaluation of the food enzyme triacylglycerol lipase from the non-genetically modified Penicillium caseifulvum strain AE-LRF.

The food enzyme triacylglycerol lipase (triacylglycerol acylhydrolase; EC 3.1.1.3) is produced with the non-genetically modified Penicillium caseifulvum strain AE-LRF by Amano Enzyme Inc. The food enzyme was free from viable cells of the production organism. It is intended to be used in four food manufacturing processes. Dietary exposure to the food enzyme-total organic solids (TOS) was estimated to be up to 0.013 mg TOS/kg body weight (bw) per day in European populations. Genotoxicity tests did not indicate a safety concern. The systemic toxicity was assessed by means of a repeated dose 90-day oral toxicity study in rats. The Panel identified a no observed adverse effect level of 69 mg TOS/kg bw per day, the highest dose tested, which when compared with the estimated dietary exposure, resulted in a margin of exposure of at least 5308. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. However, the Panel noted that traces of ■■■■■, used in the manufacture of the triacylglycerol lipase, may be found in the food enzyme. The Panel considered that the risk of allergic reactions upon dietary exposure could not be excluded, particularly in individuals sensitised to fish. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns, under the intended conditions of use.

Safety evaluation of the food enzyme ß-glucosidase from the non-genetically modified Penicillium guanacastense strain AE-GLY.

The food enzyme ß-glucosidase (ß-D-glucoside glucohydrolase; EC 3.2.1.21) is produced with the non-genetically modified Penicillium guanacastense strain AE-GLY by Amano Enzyme Inc. The food enzyme is intended to be used in four food manufacturing processes. Dietary exposure to the food enzyme-total organic solids (TOS) was estimated to be up to 4.054 mg TOS/kg body weight (bw) per day in European populations. Genotoxicity tests did not raise a safety concern. The systemic toxicity was assessed by means of a repeated dose 90-day oral toxicity study in rats. The Panel identified a no observed adverse effect level of 943 mg TOS/kg bw per day, the highest dose tested, which when compared with the estimated dietary exposure, resulted in a margin of exposure of at least 233. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.